CAR/PXR provide directives for Cyp3a41 gene regulation differently from Cyp3a11.

This study reports that Cyp3a41 gene contains 13 exons and is localized on the chromosome 5. CYP3A41 is a female-specific isoform that is predominantly expressed in the liver. Estrogen signaling is not responsible for its female specificity. CYP3A41 expression in kidney and brain is observed only in 50% of mice examined. PXR mediates dexamethasone-dependent suppression of CYP3A41. In contrast to CYP3A11, CYP3A41 expression is not induced by pregnenolone-16alpha-carbonitrile (PCN) in wild-type mice, but is significantly suppressed by PCN in PXR(-/-) mice. Phenobarbital and TCPOBOP induce CYP3A11 expression only in the presence of CAR, but have no effect on CYP3A41 expression. Immunoblot and erythromycin demethylase activity analysis reveal robust CYP3A induction after PCN treatment, which is poorly correlated to CYP3A41. These findings suggest a differential role for CAR/PXR in regulating individual CYP3A isoforms by previously characterized CYP3A inducers.

Evaluation of comparative cytochrome P450 2B4 model by photoaffinity labeling.

A homology model of rabbit CYP 2B4 was constructed on the basis of the crystallographic structure of truncated mammalian CYP 2C5/3 and bacterial soluble CYPs. To validate the CYP 2B4 homology model photoaffinity labeling was employed. Three probes (I-III) containing a photo-labile azido-group and an amino-group on opposite ends of the molecule were designed for photoaffinity labeling of the CYP 2B4 in increasing distance from the heme iron. Spectroscopic data proved probes I (the shortest) and II (a middle sized) to be coordinated with the heme iron via their amino-groups in the enzyme active center while the probe III (the longest) was not bound in this way. This binding orientation of probes I and II is in accordance with the model predicting ion-pairing of the negatively charged side chain of CYP 2B4 Asp 105 and a positively charged nitrogen located in an appropriate position in structures of probes I and II, only. The lack of heme binding of the probe III is clear from its docking into the CYP 2B4 model since no Asp 105 ion-pairing is possible. The target of photoactivated probe II, Arg 197, in a distance of about 16.5 A from the heme iron, exactly matches the position of that amino acid residue, predicted from the CYP 2B4 homology model. Moreover, using this technique, a substrate access channel has been identified. To assess the predicted substrate-binding pocket, an interaction of a specific CYP 2B4 substrate, diamantane, was examined. In "silico" docking revealed strong binding of diamantane in an orientation allowing experimentally observed C4-hydroxylation. Our homology model of CYP 2B4 is thus consistent with experimental metabolic and photoaffinity labeling data.

Expression of cytochromes P450 4F4 and 4F5 in infection and injury models of inflammation.

Lipopolysaccharide (LPS) treatment of rats suppresses CYP 4F4 and 4F5 expression by 50 and 40%, respectively, in a direct fashion occurring in the liver. This contention is borne out by essentially parallel dose-dependent changes observed upon treatment of rat hepatocyte cultures with LPS. An alternate avenue of triggering the inflammatory cascade is traumatic brain injury by controlled cortical impact. Such injury brings about a dramatic change in the expression of CYP 4F4 and 4F5 mRNA which reaches its greatest effect 24 h after impact compared with sham-operated but uninjured controls. At time points after 24 h the expression of both isoforms increases dramatically reaching highest levels at 2 weeks post-injury. These changes in mRNA expression are mirrored by changes in protein expression. The results are consistent with the notion that immediately after injury concentrations of leukotriene and prostaglandin mediators are elevated by decreased CYP 4F concentrations. As time after injury increases those conditions reverse. Increased CYP 4F expression leads to diminished concentrations of leukotriene and prostaglandin mediators and then to recovery and repair.

Bringing a family practice model of health to the People's Republic of China.

The health care system in the People's Republic of China (PRC) is undergoing a major transition that has made the government revise its approach to how medicine is taught and practiced. Family medicine, which provides a generalist approach to medical care, is at the forefront of this transition. This article reviews the recent history of medical education in the PRC, including the establishment of the discipline of family medicine in the mid 1980s, and factors promoting development of family medicine. These include the movement away from government-subsidized health care in hospital settings, the aging population, increased urbanization, increasing incidence of infectious diseases, and rising health care costs. We conclude from observations made in the PRC and from a review of secondary sources that family medicine in China is in its infancy. The value of understanding the role that family medicine plays within China's changing health care system is that we gain a broader perspective of the variety and growing international importance of family practice as a profession.

Cytochromes P450 in brain: function and significance.

The presence and activity of cytochromes P450 in brain regions and various brain cells have been extended and advanced over the last five years covered by this review. Using in situ hybridization and immunohistochemical techniques, many cytochrome P450 enzymes have been demonstrated to be present in brain and to have a regional rather than universal distribution. Many of these various cytochromes P450 have been shown to catalyze the metabolism of neurosteroids as well as other biologically significant compounds in brain. In addition, many cytochrome P450 enzymes have been implicated in the metabolism of psychoactive drugs such as neuroleptics and antidepressants. The regulation of cytochrome P450 expression has been studied at greater detail, the regulation of aromatase being a prominent example during the last five years.

Role of LYS271 and LYS279 residues in the interaction of cytochrome P4501A1 with NADPH-cytochrome P450 reductase.

It has been proposed that negatively charged amino acids on the surface of reductase and positively charged amino acids on the surface of P450 mediate the binding of both proteins through electrostatic interactions. In this study, we used a site-directed mutagenesis approach to determine a role for two lysine residues (Lys271 and Lys279) of cytochrome P4501A1 in the interaction of P4501A1 with reductase. We prepared two mutants P4501A1Ile271 and P4501A1Ile279 with a mutation of the lysine at positions 271 and 279, respectively. We observed a strong inhibition (>80%) of the 7-ethoxycoumarin and ethoxyresorufin deethylation activity in the reductase-supported system for both mutants. In the cumene hydroperoxide-supported system, P4501A1Ile279 exhibited wild-type activity, but the P4501A1Ile271 mutant activity remained low. The CD spectrum and substrate-binding assay indicated that the secondary structure of P4501A1Ile271 is perturbed. To evaluate further the involvement of these P4501A1 lysine residues in reductase binding, we measured the KM of reductase for wild type and mutants. Both wild type and P4501A1Ile271 reached saturation in the range of reductase concentrations tested with KM values 5.1 and 11.2 pM, respectively. The calculated KM value for P4501A1Ile279 increased 9-fold, 44.4 pM, suggesting that the mutation affected binding of reductase to P4501A1. Stopped-flow spectroscopy was employed to evaluate the effect of mutations on electron transfer from reductase to heme iron. Both wild type and P450Ile279 showed biphasic kinetics with a approximately 40% participation of the fast step in the total activity. On the other hand, only single-phase kinetics for iron reduction was observed for P450Ile271, suggesting that the low activity of this mutant can be attributed not only to major structural changes but also to a disturbance in the electron transport.

Role of THR501 residue in substrate binding and catalytic activity of cytochrome P4501A1.

A putative binding region for cumene hydroperoxide in the active site of cytochrome P4501A1 was identified using photoaffinity labeling. Thr501 was determined as the most likely site of modification by azidocumene used as the photoaffinity label (T. Cvrk and H. W. Strobel, (1998) Arch. Biochem. Biophys. 349, 95-104). To evaluate further the role of this amino acid residue a site-directed mutagenesis approach was employed. P4501A1 wild type and two mutants, P4501A1Glu501 and P4501A1Phe501, were expressed in and purified from Escherichia coli and used for kinetic analysis to confirm the role of Thr501 residue in cumene hydroperoxide binding. The mutation resulted in a two- to fourfold decrease in the rate of heme degradation in the presence of 0.5 mM cumene hydroperoxide. The mutations do not prevent or significantly alter binding of the tested substrates; however, binding of 2-phenyl-2-propanol (product generated from cumene hydroperoxide) to P4501A1Glu501 and P4501A1Phe501 exhibited four- and eightfold decreases, respectively, suggesting that the mutations strongly affected the affinity of cumene hydroperoxide for the P4501A1 active site. The kinetic analysis of cumene hydroperoxide-supported reactions showed that both mutants exhibit increased Km and decreased VMax values for all tested substrates. Furthermore, the mutations affected product distribution in testosterone hydroxylation. On the basis of P4501A1Glu501 and P4501A1Phe501 characterization, it can be concluded that Thr501 plays an important role in cumene hydroperoxide/P4501A1 interaction.

Molecular cloning and regulation of expression of two novel mouse CYP4F genes: expression in peroxisome proliferator-activated receptor alpha-deficient mice upon lipopolysaccharide and clofibrate challenges.

Cytochrome P450 4F isoforms catalyze the hydroxylation of eicosanoids such as leukotriene B(4), prostaglandins, and lipoxins as well as hydroxyeicosatetraenoic acids. In the present study, we report the molecular cloning of two novel mouse CYP4F isoforms, CYP4F15 and CYP4F16. Sequence comparison showed that CYP4F15 has 93.5% homology to CYP4F4 and CYP4F16 has 90.8% homology to CYP4F5, therefore they are the orthologs for rat CYP4F4 and CYP4F5, respectively. Both isoforms are expressed in liver and also in extrahepatic tissues but the patterns of expression are slightly different. To elucidate further the regulation and regulatory mechanism of the two isoforms, renal and hepatic CYP4F15 and CYP4F16 expression were analyzed using wild-type (SV/129) mice and peroxisome proliferator-activated receptor (PPAR) alpha null mice with or without challenge by bacterial endotoxin (LPS) or clofibrate. Renal expression of CYP4F15 was induced by LPS and clofibrate in (+/+) mice, and these effects were absent in the (-/-) mice. Renal expression of CYP4F16 was not affected by LPS or clofibrate in (+/+) or (-/-) mice. In contrast, hepatic expression of CYP4F15 and CYP4F16 was significantly reduced by LPS-treatment in (+/+) mice. A lesser reduction was also seen in the (-/-) mice, suggesting that PPARalpha is partially responsible for this down-regulation. Clofibrate treatment caused the reduction of hepatic CYP4F16 expression and this effect was not dependent on PPARalpha. Clofibrate treatment had no effect on hepatic CYP4F15 expression. Together, our data indicate that CYP4Fs are regulated in an isoform-specific, tissue-specific, and species-specific manner.

In vitro metabolism of chlorpromazine by cytochromes P450 4F4 and 4F5 and the inhibitory effect of imipramine.

The metabolism of chlorpromazine by expressed recombinant cytochromes P450 4F4 and 4F5 cloned from rat brain was analyzed to characterize the individual activities of the isoforms. Both isoforms metabolized chlorpromazine to both the N-demethylated and the S-oxide products. When isoforms were incubated with chlorpromazine in the presence of increasing concentrations of imipramine, imipramine significantly inhibited both N-demethylation and S-oxidation of chlorpromazine. A dilution of the serum fraction of anti-4F antibody was also found to significantly inhibit both S-oxidation and N-demethylation of chlorpromazine by both 4F4 and 4F5.

Cytochrome P450 3A9 catalyzes the metabolism of progesterone and other steroid hormones.

The catalytic requirements and the role of P450 3A9, a female-specific isoform of CYP3A from rat brain, in the metabolism of several steroid hormones were studied using recombinant P450 3A9 protein. The optimal steroid hormone hydroxylase activities of P450 3A9 required cholate but not cytochrome b5. P450 3A9 was active in the hydroxylation reactions of testosterone, androstenedione, progesterone and dehydroepiandrosterone (DHEA). No activity of P450 3A9 toward cortisol was detectable under our reconstitution conditions. Among all the steroid hormones examined, female-specific P450 3A9 seemed to catalyze most efficiently the metabolism of progesterone, one of the major female hormones, to form three mono-hydroxylated products, 6beta-, 16alpha-, and 21-hydroxyprogesterone. Our data also showed that P450 3A9 can catalyze the formation of a dihydroxy product, 4-pregnen-6beta, 21-diol-3, 20-dione, from progesterone with a turnover number, 1.3 nmol/min/nmol P450. Based on the Vmax/Km values for P450 3A9 using either 21-hydroxprogesterone or 6beta-hydroxyprogesterone as a substrate, 4-pregnen-6beta, 21-diol-3, 20-dione may be formed either by 6beta-hydroxylation of 21-hydroxprogesterone or 21-hydroxylation of 6beta-hydroxyprogesterone. As a major isoform of CYP3A expressed in rat brain, the activities of P450 3A9 toward two major neurosteroids, progesterone and DHEA suggested a possible role for P450 3A9 in the metabolism of neurosteroids.

Chronic intragastric infusion of ethanol-containing diets induces CYP3A9 while decreasing CYP3A2 in male rats.

The CYP3A subfamily is the most abundant of the human hepatic cytochrome P450 enzymes. They mediate the biotransformation of many drugs, including a number of psychotropic, cardiac, analgesic, hormonal, immunosuppressant, antineoplastic, and antihistaminic agents. We studied diet/ethanol interactions using total enteral nutrition in adult male Sprague-Dawley rats with diets containing 16% protein, ethanol (13 g/kg), corn oil (fat; 25-45%), and carbohydrate (CHO; 1-21%). Using this model, chronic ethanol feeding decreased CYP3A activity (testosterone 6 beta-hydroxylation) and apoprotein levels (Western blot) (P <.05) and these effects were independent of the dietary CHO/fat ratio. The CYP3A2 mRNA levels decreased (P <.05) in the rats fed ethanol-containing diets by 73 to 83% compared with rats fed control diets, regardless of the CHO/fat ratio. In contrast, ethanol induced CYP3A9 mRNA levels (P <.05) and this effect was greater (P <.05) in the high-CHO/low-fat group (11.3-fold) than in the low-CHO/high-fat group (2.6-fold). Purified recombinant rat P450 3A9 had a chlorzoxazone 6-hydroxylase activity with a turnover number 1.3 nmol/min/nmol of P450. These results indicate that 1) ethanol differentially affects the expression of CYP3A gene family and this regulation appears to be modulated by dietary CHO/fat ratio; 2) the decrease in testosterone 6 beta-hydroxylase activity and CYP3A apoprotein levels are most likely due to the ethanol-induced decrease in CYP3A2 mRNA levels; and 3) CYP3A9 is induced by ethanol and is a low-affinity, high-K(m) chlorzoxazone hydroxylase.

Recombinant cytochrome P450 2D18 metabolism of dopamine and arachidonic acid.

The function of cytochrome P450 (P450) in the mammalian brain is not well understood. In an effort to further this understanding, this study identifies two endogenous substrates for P450 2D18. Previous reports have shown that this isoform is expressed in the rat brain, and the recombinant enzyme catalyzes the N-demethylation of the antidepressants imipramine and desipramine. By further examining the substrate profile of P450 2D18, inferences can be made as to potential endogenous P450 substrates. Herein we demonstrate the metabolism of the central nervous system-acting compounds chlorpromazine and chlorzoxazone with turnover numbers of 1.8 and 0. 9 nmol/min/nmol, respectively. Because the four aforementioned pharmaceutical substrates work by binding to neurotransmitter receptors, binding assays and oxidation reactions were performed to test whether dopamine is a substrate for P450 2D18. These data indicate a K(S) value of 678 microM and that P450 2D18 can support the oxidation of dopamine to aminochrome through a peroxide-shunt mechanism. We also report the P450 2D18-mediated omega-hydroxylation and epoxygenation of arachidonic acid, primarily leading to the formation of 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acids, compounds that have been shown to have vasoactive properties in brain, kidney, and heart tissues. The data presented herein suggest a possible role for P450 involvement in membrane and receptor regulation via epoxyeicosatrienoic acid formation and a potential involvement of P450 in the oxidation of dopamine to reactive oxygen species under aberrant physiological conditions where the sequestering of dopamine becomes compromised, such as in Parkinson's disease.

A novel human cytochrome P450 4F isoform (CYP4F11): cDNA cloning, expression, and genomic structural characterization.

By a combination of cDNA library screening and rapid amplification of cDNA ends analysis, a novel human cytochrome P450 4F isoform has been cloned and sequenced. The new 4F isoform is designated CYP4F11 and contains 1765 nucleotides. The coding region encodes 524 amino acid residues, and the heme-binding region is highly conserved. The CYP4F11 amino acid sequence has 80.0, 82.3, and 79.2% identity to CYP4F2, CYP4F3, and CYP4F8 amino acid sequences, respectively. In vitro translation shows the molecular mass of CYP4F11 is approximately 57 kDa, consistent with the calculated molecular mass. CYP4F11 is expressed mainly in human liver, followed by kidney, heart, and skeletal muscle. The genomic structure of CYP4F11 was solved by database searching and computer analysis. The coding region of CYP4F11 has 12 exons. The CYP4F11 gene is located 16 kb upstream of the CYP4F2 gene on chromosome 19. This is consistent with the notion that the human cytochrome P450 4F genes form a cluster on chromosome 19.

Cytochrome P-450 activities in human and rat brain microsomes.

The role of cytochrome P450 in the metabolism of dextromethorphan, amitriptyline, midazolam, S-mephenytoin, citalopram, fluoxetine and sertraline was investigated in rat and human brain microsomes. Depending on the parameters, the limit of quantification using gas chromatography-mass spectrometry methods was between 1.6 and 20 pmol per incubation, which generally contained 1500 microg protein. Amitriptyline was shown to be demethylated to nortriptyline by both rat and human microsomes. Inhibition studies using ketoconazole, furafylline, sulfaphenazole, omeprazole and quinidine suggested that CYP3A4 is the isoform responsible for this reaction whereas CYP1A2, CYP2C9, CYP2C19 and CYP2D6 do not seem to be involved. This result was confirmed by using a monoclonal antibody against CYP3A4. Dextromethorphan was metabolized to dextrorphan in rat brain microsomes and was inhibited by quinidine and by a polyclonal antibody against CYP2D6. Only the addition of exogenous reductase allowed the measurement of this activity in human brain microsomes. Metabolites of the other substrates could not be detected, possibly due to an insufficiently sensitive method. It is concluded that cytochrome P450 activity in the brain is very low, but that psychotropic drugs could undergo a local cerebral metabolism which could have pharmacological and/or toxicological consequences.

Heterobifunctional photoaffinity probes for cytochrome P450 2B.

Three heterobifunctional photoaffinity probes, N-(p-azidobenzyl)-N-methyl-p-aminobenzylamine (I), N-(p-azidobenzyl)-N-methyl-p-aminophenethylamine (II), and N-(p-azidophenethyl)-N-methyl-p-aminophenethylamine (III), were synthesized and characterized. These probes, containing a photolabile azido-group and an amino-group on opposite sides of the molecule, were designed for photoaffinty labeling of the cytochrome P450 (CYP) 2B active site cavity differing in distance from the heme iron. Spectroscopic studies proved that probes I and II coordinated with the heme iron via their amino-group in the enzyme active center, whereas probe III did not. This result in conjunction with data from kinetic studies suggests probes I and II are appropriate for photoaffinity labeling of the CYP 2B active center. Thus, probe II was used to identify amino acid residues within a distance of the probe length (about 16.5 A) from the heme. Analysis of a Lys-C digest of the probe II-labeled CYP 2B4 revealed a single labeled hexapeptide corresponding to position 192-197 of the CYP 2B4 sequence. Using postsource decay/matrix-assisted laser desorption ionization-time of flight, Arg197 was identified as a probe II target. The location of the labeled site in three-dimensional structures of bacterial CYPs and in CYP 2B homology models is discussed.

Extrahepatic drug metabolizing enzymes.

Drug metabolizing enzymes are known to be present and active in most extrahepatic tissues. In this review, evidence is presented that expression and activity of several extrahepatic drug-metabolizing enzymes are regulated in unique ways, which may be associated with tissue functions and activities. In several instances evidence is offered that hormonal effects may be regulated through tissue specific distribution and/or responses of transcription factors.

Metabolism of cyclosporine by cytochromes P450 3A9 and 3A4.

The ability of P450 3A9 to transform cyclosporine was studied and compared to that of human P450 3A4. Purified P450 3A4 and P450 3A9 proteins were reconstituted in a system containing potassium phosphate buffer, lipids, NADPH-P450 reductase, and glutathione with NADPH added to initiate the reaction. Cyclosporine was added alone and with or without the inhibitors, ketoconazole or troleandomycin. High performance liquid chromatography with ultraviolet (HPLC/UV) techniques were used to analyze for cyclosporine metabolites. Both P450 3A4 and P450 3A9 transformed cyclosporine to three metabolites: AM1, AM9, and AM4n. P450 3A4 predominantly formed AM1 (63% of metabolites formed) while P450 3A9 formed AM4n (59% of metabolites formed). Ketoconazole (0.5 microM) completely inhibited P450 3A9 catalyzed formation of AM1 and AM9 and reduced AM4n formation to 28% of control. AM4n, AM1, and AM9 formation catalyzed by P450 3A4 was reduced to 50%, 30%, and 10% of control, respectively, by 0.5 microM ketoconazole. Troleandomycin (> 10 microM) inhibited the formation of AM4n by P450 3A4 and P450 3A9 to 60-70% of control, while the production of AM1 by P450 3A4 was increased to 120% of control and the production of AM1 by P450 3A9 was inhibited to 50% of control. Inhibition of P450 3A4 by troleandomycin (> 10 microM) reduced the formation of AM9 to 40% of control, but only reduced P450 3A9 formation of AM9 to 80% of control. This study shows that rat P450 3A9 is capable of transforming cyclosporine to multiple metabolites similar to those generated by human P450 3A4.

High-performance liquid chromatographic methods for the analysis of haloperidol and chlorpromazine metabolism in vitro by purified cytochrome P450 isoforms.

Sensitive HPLC methods for the resolution and quantitation of metabolites of both haloperidol and chlorpromazine metabolism have been developed for use in in vitro reconstitution assays with purified P450 isoforms. Separation of haloperidol metabolites was accomplished using a Hypersil CPS column with a mobile phase of 67% acetonitrile and 10 mM ammonium acetate, pH 5.4. Separation of chlorpromazine metabolites was achieved using an Ultrasphere cyano column with a mobile phase of 87.5% acetonitrile, 5% methanol, 3% 0.12 M sodium acetate, 3% 0.12 M ammonium acetate, 0.01% diethylamine and 0.01% triethylamine, pH 9.5. Sharp resolution was observed for haloperidol and three of its major metabolites and for chlorpromazine and five of its major metabolites. Varying levels and combinations of these metabolites are formed during in vitro incubations of parent compound with purified P450 isoforms 1A1 and 2B1 in a reconstituted system.

Isolation of partially purified P450 2D18 and characterization of activity toward the tricyclic antidepressants imipramine and desipramine.

Previous reports have shown that rat brain microsomes are capable of metabolizing tricyclic antidepressants such as imipramine. Subsequent studies have shown that the protein products of several clones isolated from rat brain cDNA libraries are capable of metabolizing imipramine to both its active metabolite, desipramine, and its inactive hydroxylated metabolites. We report here the overexpression and partial purification of P450 2D18 using the baculovirus expression system and the incorporation of a C-terminal [His]4 tag. P450 2D18 was partially purified to a specific content of 4.8 nmol/mg protein and shown to be electrophoretically pure. The apparent KM values for P450 2D18 toward imipramine and desipramine were 374 and 314 microM, respectively. While apparent KM values were similar, P450 2D18 was shown to have a fivefold increased Vmax (2.2 nmol/min/nmol P450) for imipramine compared to desipramine (0.44 nmol/min/nmol P450), suggesting a primary involvement in the activation of imipramine to desipramine. We also examined the effect of the CYP2D6 inhibitor quinidine, the CYP3A inhibitor ketoconazole, and the dopamine reuptake inhibitor GBR-12935 for their ability to inhibit P450 2D18-mediated metabolism of imipramine. These results, when compared with previous studies using rat brain microsomes, suggest that P450 2D18 may play an important role in the conversion of imipramine to its active metabolite desipramine in the rat brain.

Expression, induction and regulation of the cytochrome P450 monooxygenase system in the rat glioma C6 cell line.

The cytochrome P450 monooxygenase system consists of NADPH-cytochrome P450 reductase (P450 reductase) and cytochromes P450, which can catalyze the oxidation of a wide variety of endogenous and exogenous compounds. P450 reductase transfers reducing equivalents from NADPH to P450, which in turn catalyzes metabolic reactions. In previous studies, we have used the rat glioma C6 cell line as an in vitro model system and identified the presence of P450 reductase and of cytochrome P450 1A1, 1A2, 2A1, 2B1/2, 2C7, 2D1-5 and 2E1 by reverse transcription followed by polymerase chain reaction (RT-PCR). In C6 cells, the induction of P450 1A1 and 2B1/2 at mRNA level after BA (benzo(a)anthracene) or PB (phenobarbital) treatments was detected. In this study, analysis of microsomal preparations of glioma C6 cells was utilized to demonstrate the presence of P450 2B and P450 reductase at the protein level. ELISAs showed that PB induced P450 2B proteins 12-fold. These experiments further establish that the rat glioma C6 cell line contains an active cytochrome P450 monooxygenase system that can be induced by P450 inducers. We also found that the mRNAs of P450 1A1 and 2B1/2 from glioma C6 cells do not bind to the oligo(dT)-based separation techniques efficiently, suggesting that they may have very short poly(A) tails. The half-lives of P450 1A1 and 2B1/2 mRNA in glioma C6 cells are 1/10 and 1/3 of that in liver, respectively. This may partly contribute to the low expression level of P450s in glial cells. The induction of P450s by BA or PB did not change their mRNA half-lives, indicating the induction may be due to transcriptional regulation. In summary of this study, we believe the presence of the cytochrome P450 monooxygenase system in glial cells of the brain may be important in chemotherapy and carcinogenesis of brain tumors.